INHIBITION OF PEPTIDE SIGNAL TRANSDUCTION--THERAPY & PREVENTION OF LUNG CANCER

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The overall objective of this project is to develop new agents and routes of delivery for the chemoprevention and treatment of human lung cancer. We propose to do this by developing agents which interfere with aberrant signal pathways in human lung cancer and dysplastic bronchial epithelial cells. We will deliver these agents by transtracheal instillation and aerosolized inhalation in a preclinical model using athymic rats bearing orthotropic human lung cancers. We propose a stepwise approach which begins with in vitro studies in human lung cancer and bronchial epithelial cell lines. These studies are followed by normal animal studies to evaluate toxicity, delivery and pharmacokinetic parameters and orthotopic animal studies to evaluate tumor delivery and response. Phase I and II human clinical studies re athe last step in the evaluation process. Lung cancer is the most lethal cancer in the United States and the cure remains low at 13% primarily because th cancers have metastasized at the time of diagnosis. Thus, new early detection, prevention and treatment strategies are needed. Many lung cancers (all small cell lung cancers and some non- small cell lung cancers) have a neuroendocrine phenotype. These cells express multiple neuropeptide receptors and neuropeptides activate an intracellular signal pathway in these cells. This pathway is activated by peptide-receptor binding. The receptors are linked to heterotrimeric G proteins ( of the alphaGq family and beta and gamma) which when activate in turn activate phospholipase Cbeta (PLCbeta). Activated PLCbeta cleaves membrane lipids to inositol trisphosphate (IP3) which binds to receptors on the endoplasmic reticulum resulting in a rise intracellular calcium. The PLCbeta activation also results in activation of protein kinase C through diacylglycerol. The free calcium activates cytoplasmic protein which subsequently act as transcription factors leading to cell proliferation. During the past SPORE grant period, we shoed that peptide activation of G proteins can also lead to activation of the MEKK pathway. Alteration of the signal pathway by constitutively active GTPase deficient Gq protein genes introduced in a retrovirus or by treatment with substance p derivatives led to discordant signaling. The discordant signaling was characterized by inhibition of PLCbeta activation, inhibition of calcium release and inhibition of proliferation with stimulation of the MEKK pathway and induction of apoptosis. We also showed that a series of new dimeric bradykinin antagonists also inhibited calcium release and inhibited growth while inducing apoptosis. We began to explore the delivery of these genetic and pharmacologic agents via transtracheal instillation and aerosol inhalation in an orthotopic nude mouse and rat model. We also showed that a novel PKC/calmodulin inhibitor, dexniguldipine, inhibited SCLC growth in vitro and in vivo, determined a dose the phase II studies by completing a phase I study and have accrued 14 patients to a phase I study. During the next 5 years, we will complete this phase II dexniguldipine study; complete in vitro evaluation of G protein genes and antisense oligomers delivered in polycationic lipids and human retrovirus vectors; optimize in vivo delivery of antisense oligonucleotides and mutant cell signal genes in polycationic lipid and retroviral vectors in nude rats bearing orthotopic lung cancers after delivery by intratracheal instillation and aerosolized inhalation; and determine maximally effective new substance P and bradykinin antagonists. We believe that these new treatment and prevention strategies will be applied to human subjects in the next 5 years and will ultimately lead to advances in lung cancer mortality.